The value of electrophoresis in clinical chemistry has been recognized for some time in the analysis of proteins in body fluids. Gel electrophoresis is the older method used in clinical chemistry laboratories. With gel electrophoresis, a sample is applied near one edge of a layer of gelatin carried on a flexible sheet slab of gel. The gel is electrophoresised, stained and the density of the resulting pattern is measured to reveal the sample such as proteins and nucleic acids. Although gel electrophoresis is relatively inexpensive in terms of the supplies and equipment required to perform sample analyses, the technique requires skilled technicians and is time consuming, effectively resulting in a high price per test and limiting the number of tests that can be performed using the technique.
Efforts have been made to improve electrophoresis in the clinical laboratory. For example, U.S. Pat. No. 4,124,470 to Dahms describes a zone electrophoresis apparatus where a number of samples in individual large-bore tubes can be processed serially on a turntable.
Capillary electrophoresis is a more recent development. In capillary electrophoresis, a small tube or capillary having an inside bore diameter in the range of about five microns to about two hundred microns and often about twenty cm long is filled with an electrically conductive fluid, or buffer. A direct current voltage in a range of about 2,000 volts to about 30,000 volts is applied to the ends of the capillary by means of electrodes positioned in the buffer reservoirs, causing a small current, typically in the range of about five microamps to about one milliamp, to flow through the capillary.
With the correct polarity applied across the capillary, the sample begins to migrate from the sample introduction end toward the other end of the capillary. As this migration occurs, different molecules in the sample travel at different rates primarily because of slightly different electrical charges on the molecules. These different migration rates cause molecules with slightly different charges to separate one from the other, some moving more quickly and advancing relatively with respect to more slowly moving molecules. As the sample nears the other end of the capillary, the small volume of sample becomes separated into bands of different molecules according to the relative migration rates of the molecules. These bands or groups of different molecules are detected near the other end of the capillary by, for example, passing a light beam through the bore of the capillary. Changes to the light beam, such as absorbance caused by the different molecules, are detected as the separated molecules pass through the beam, thus identifying the different molecules or the classes or categories of molecules in the sample and the relative concentration of such molecules.
The core of each capillary electrophoresis system is the capillary. Although there are capillaries made of borosilicate glass or Teflon, fused silica is by far the most frequently used material. This is owing to the intrinsic properties of fused silica like the superior transparency for UV light, the high thermal conductance and the feasibility of manufacturing capillaries with diameters of a few micrometers. Because bare fused silica capillaries are extremely fragile, they are externally coated with a polyimide polymer to improve the flexibility and to make handling easier. However, a small amount of the polyimide coating has to be removed to allow light passage for detection.
Capillary electrophoresis analyzers using small bore tubes are known in the art. For example, in European Patent Application Number 89302489.3, Publication Number 0,339,779 A2, the ends of the capillaries along with electrodes are inserted into the vials by means of hypodermics that pierce caps on the vials. A single detector provides detection of electrophoresised samples.
Another automated capillary electrophoresis apparatus is described in U.S. Pat. No. 5,045,172 to Guzman. A capillary, which is described in Guzman as being a single capillary or a plurality of capillary tubes operated in parallel or in a bundle, has two opposite ends. Other small bore tube capillary electrophoresis devices are described in Klein, U.S. Pat. No. 5,413,686 and Waska et al, U.S. Pat. No. 5,312,535.
As an alternative to the cylindrical capillary glass tubes, rectangular capillaries have been disclosed by Tsuda et al, "Rectangular Capillaries for Capillary Zone Electrophoresis", Anal. Chem. 62 (1990) 2149-2152. Some of the advantages of these systems are their efficient heat dissipation due to the large height-to-width ratio and, hence, their high surface-to-volume ratio and their high detection sensitivity for optical on-column detection modes. According to the authors, these flat separation channels have the ability to perform two-dimensional separations, with one force being applied across the separation channel, and with the sample zones detected by the use of a multi-channel array detector. A significant disadvantage of rectangular capillaries are their thin walls, equal to the width of the narrowest dimension; for instance 20 um for 20.times.20 um.sup.2 capillaries. Special care must be taken to avoid breakage of these capillaries. The rectangular as well as circular capillaries of the prior art do not allow multiple electrodes or detectors at any one or more desired points between the ends of the fluid pathway.
The present invention provides a novel microfluidic device useful in capillary electrophoresis analyzers and other fluidic devices for separation and analysis of proteins, nucleic acids and other chemical compounds.